The research outlined in this proposal will investigate the growth inhibitory response of epithelial cells to Transforming Growth Factors-beta (TGF-beta). Changes in epithelial cell responsiveness to TGF-beta has been associated with the carcinogenic process of a large number of epithelial cancers. Further genetic evidence suggests that tumor suppressor genes are inactivated as part of the development of epithelial cancers. Since many tumor suppressor genes are growth inhibitory in the normal physiology of cells, it is possible that some elements of the TGF-beta growth inhibitory pathway of epithelial cells are tumor suppressor genes. The model system used is the mink lung epithelial cell line (CCL 64) in which the investigator has generated a panel of mutants defective in TGF-beta response- in some cases because of biochemically identifiable defects in the TGF-beta receptor. In addition, the investigator has developed a unique in vitro assay for the identification of growth inhibitory cDNAs. This assay is based on the dilution of a lipophilic fluorescent dye among daughter cells as cells divide. If a cell is transfected with a cDNA which slows the transfectants doubling time, the progeny of that cell will retain more dye over time. The population is allowed to grow for approximately four population doublings at which time the brightest cells in the population are isolated and cDNAs associated with that slowly growing population are identified. TGF-beta's growth-inhibitory effects will be investigated using this system by constructing cDNA libraries which are enriched for messages regulated by TGF-beta, transfecting these libraries into TGF-beta-nonresponsive Mv1Lu mutants, and identifying cDNAs induced by TGF-beta which can slow cells growth rates. These TGF-beta-regulated growth inhibitory genes will subsequently be screened for tumor suppressor activity by a variety of genetic and physiological criteria.